Methylthioadenosine deaminase in an alternative quorum sensing pathway in Pseudomonas aeruginosa

Pseudomonas aeruginosa possesses an unusual pathway for 5′-methylthioadenosine (MTA) metabolism involving deamination to 5′-methylthioinosine (MTI) followed by N-ribosyl phosphorolysis to hypoxanthine and 5-methylthio-α-d-ribose 1-phosphate. The specific MTI phosphorylase of P. aeruginosa has been reported [Guan, R., Ho, M. C., Almo, S. C., and Schramm, V. L. (2011) Biochemistry 50, 1247-1254], and here we characterize MTA deaminase from P. aeruginosa (PaMTADA). Genomic analysis indicated the PA3170 locus to be a candidate for MTA deaminase (MTADA). Protein encoded by PA3170 was expressed and shown to deaminate MTA with 40-fold greater catalytic efficiency for MTA than for adenosine. The k_cat/K _m value of 1.6 × 10⁷ M^-1 s^-1 for MTA is the highest catalytic efficiency known for an MTA deaminase. 5′-Methylthiocoformycin (MTCF) is a 4.8 pM transition state analogue for PaMTADA but causes no significant inhibition of human adenosine deaminase or MTA phosphorylase. MTCF is permeable to P. aeruginosa and exhibits an IC ₅₀ of 3 nM on cellular PaMTADA activity. PaMTADA is the only activity in P. aeruginosa extracts to act on MTA. MTA and 5-methylthio-α-d-ribose are involved in quorum sensing pathways; thus, PaMTADA is a potential target for quorum sensing. The crystal structure of PaMTADA in complex with MTCF shows the transition state mimic 8(R)-hydroxyl group in contact with a catalytic site Zn²⁺, the 5′-methylthio group in a hydrophobic pocket, and the transition state mimic of the diazepine ring in contact with a catalytic site Glu.